The dorsomedial nucleus of the hypothalamus (DMH) is a major relay and integrator in the circuitry through which circadian patterns in arousal state, behavior and endocrine rhythms are temporally organized. Cell-specific lesions of the DMH produce a significant reduction in the amplitude of circadian rhythms of locomotor activity, wakefulness, corticosteroid secretion and feeding. Although it is clear that the DMH occupies a prominent role in the circuitry connecting the circadian timing system to effector circuits driving arousal and endocrine rhythms, it is not known which neurotransmitter(s) it releases to drive downstream targets. It has been proposed that excitatory glutamatergic inputs from the DMH to wake-promoting neurons of the lateral hypothalamus (LH) and inhibitory GABAergic inputs from the DMH to sleep-promoting neurons in the ventrolateral pre-optic area (VLPO) may be critical for circadian modulation of sleep, activity, and feeding rhythms. Circadian regulation of corticosteroid release may depend on an excitatory projection from the DMH to corticotropin releasing hormone-expressing neurons in the paraventricular hypothalamus (PVH). Evidence for the above hypothesis comes from neuroanatomical and physiological studies in the rat. We aim to test the extent to which glutamatergic and GABAergic outputs of the DMH drive circadian oscillation in sleep, locomotor activity, feeding and corticosteroid release by selectively preventing the release of glutamate or GABA from only DMH neurons in conditional knock out mice and recording effects on all the relevant physiological and behavioral outputs. Aim 1: To compare the anatomy and neurotransmitters of DMH efferents in the mouse with rat - using anterograde tracing from the DMH and retrograde tracing from the LH, PVH and VLPO, coupled with immunohistochemistry and in situ hybridization for appropriate markers. Aim 2: To assess the role of glutamate in DMH modulation of circadian rhythms in activity, sleep, feeding, Tb and corticosterone release - by injecting an adeno-associated viral vector expressing Cre- recombinase into the DMH of conditional knock-out mice bearing Lox-P within the gene for a critical vesicular glutamate transporter Aim 3: To assess the role of GABA in DMH modulation of circadian rhythms in activity, sleep, feeding, Tb and corticosterone release - by injecting an adeno-associated viral vector expressing Cre- recombinase into the DMH of conditional knock-out mice bearing Lox-P within the gene for a critical vesicular GABA transporter. PUBLIC HEALTH RELEVANCE: Our project focuses on elucidating neural circuits that drive daily rhythms in wakefulness, activity, feeding, and hormone secretion and keep these functions synchronized. There is evidence that disruption of the synchrony between these functions can substantially impair sleep quality (1), diminish cognitive performance (2), and is a risk factor for mental health/affect disorders (3, 4), hypertension (5), diabetes and metabolic deficits (3, 6, 7), cancer (8, 9), and higher mortality rate (10). It is our hope that by understanding the neural circuitry underlying temporal synchronization of physiology and behavior we may reveal specific risk factors and mechanisms for prevention and treatment of some of these diseases or disorders.